The stated objective of this proposal is to elucidate the components and mechanisms involved in the posttranslational processing of the critical integral membrane protein, dystroglycan. Dystroglycan is a member of the dystrophin glycoprotein complex that serves, in part, as a link between the extracellular matrix and actin cytoskeleton in many cell types. Dystroglycan requires extensive posttranslational modification including proteolytic cleavage and glycosylation for function. Defects in dystroglycan posttranslational processing can result in severe forms of muscular dystrophy. Thus far six genes have been associated with dystroglycan glycosylation and human congenital muscular dystrophies. Much remains unclear on the precise function of each identified gene and their contribution to dystroglycan processing. Current evidence also suggests that additional modifiers of dystroglycan remain unidentified. This proposal challenges the hypothesis that proper regulation of positive posttranslational pathway effectors and negative pathway regulators are required for functional dystroglycan posttranslational processing. The proposed research utilizes mouse fibroblast culture as a means to manipulate dystroglycan posttranslational processing by RNA interference. Through comprehensive analysis of knockdown cultures of each known dystroglycan processing component, the proposal will address their relative contributions and activity thresholds in normal dystroglycan processing. In this way we will better understand the etiology of dystroglycan modification related muscular dystrophies. Next, the research will identify novel mechanisms critical to dystroglycan processing via independent mouse genome knockdown library genetic screens on the basis of epitope recognition and old-world arenovirus susceptibility. These unbiased approaches will uncover required components involved in dystroglycan processing, offer new candidate genes for involvement in muscular dystrophies and novel therapeutic strategies. Congenital muscular dystrophies result in extreme muscle weakness and can include brain and eye developmental abnormalities. These diseases are a result of improper modification of the critical cell membrane protein, dystroglycan. This research seeks to better understand the mechanisms of dystroglycan modification through investigation of both known components and identification of new mechanisms. Through this research we seek to improve patient diagnosis and offer potential treatment strategies.